Currently, there are over 80,000 chemicals in use and approximately 2000 new chemicals are introduced into use every year. Regulatory agencies have recognized the need for alternative toxicological methods and models to decrease the time and expense of current toxicity testing protocols. In association with the Division of the National Toxicology Program (NTP), our group is evaluating C. elegans as an alternative organism for in vivo toxicological testing. Short life cycles, easy and inexpensive maintenance and culturing, and detailed biological knowledge has allowed for the development of rapid, low-cost toxicity tests that readily lend themselves to mechanistic studies of toxicant actions. Because of the evolutionarily conserved nature of the stress-response and other relevant pathways, it is likely that responses elicited in C. elegans will be applicable to understanding similar processes in higher organisms, including humans. This group is part of the Division of the National Toxicology Program and within the Biomolecular Screening Branch. The screening facility is involved in the development of C. elegans as an alternative organism for toxicological testing. There are two major activities of this group: research and development of C. elegans medium throughput screening (MTS) and high throughput screening (HTS) technologies and participation in the memorandum of understanding (MOU) among the NTP, EPA, and NCATS, designated as Tox21 (Science 15 February 2008 319: 906-907 DOI: 10.1126/science.1154619). In FY2015, the screening facility moved into the DNTP Lab Branch, and its capabilities have expanded to include 2D and 3D models of human xenobiotic metabolism. As a result, this is the last year that this lab will be considered a core facility. Development of C. elegans MTS assays - Protocols for the monitoring of growth, size, reproduction, feeding, and movement have been developed and have been used to test over 100 toxicants. Included in the creation of monitoring protocols, statistical analysis routines have been developed specifically for the C. elegans data. We have submitted one manuscript on the feeding assay and are completing additional studies that will be included in manuscripts currently being prepared on the reproduction and growth assays published manuscripts on the development and application of the feeding, reproduction, and growth assays as well as reviews on their combined use. In collaboration with Grace Kissling (EDBP), Marjolein Smith (SRA) we have developed a mathematical model that described C. elegans growth and the effects of toxicants on growth parameters. A manuscript describing the developmental/growth effects after exposure to ToxCast Phase I and II libraries was accepted to EHP during FY2015. This year the WormTox group tested: 1) 63 compounds that were active in an HTS MMP assay at NCATS; and 2) 12 compounds associated with the Elk River spill in West Virginia. We developed a new assay to measure mitochondrial function by measuring ATP in vivo levels. Using a strain of C. elegans which constitutively expresses firefly luciferase, we track levels of ATP pools as a marker for physiological status. Changes in luminescence following toxicant exposure reflect changes in ATP levels. This assay will be used to determine the relative effects of a variety of chemicals on physiological health instead of the more common changes in morphological traits. Through a Tox21collaboration with Menghang Xia (NCATS), we screened 63 chemicals for effects on ATP levels and larval development in C. elegans.